The exocyclic groups of DNA modulate the affinity and positioning of the histone octamer

Abstract

To investigate the nature of the chemical determinants in DNA required for nonspecific binding and bending by proteins we have created a novel DNA in which inosine-5-methylcytosine and 2,6-diaminopurine-uracil base pairs are substituted for normal base pairs in a defined DNA sequence. This procedure completely switches the patterns of the base pair H bonding and attachment of exocyclic groups. We show that this DNA binds a histone octamer more tightly than normal DNA but, surprisingly, does not alter the orientation of the sequence on the surface of the protein. However, in general, the addition or removal of DNA exocyclic groups reduces or increases, respectively, the affinity for the histone octamer. The average incremental change in binding energy for a single exocyclic group is Ϸ40 J͞mol. The orientation of the DNA in core nucleosomes also is sensitive to the number and nature of the exocyclic groups present. Notably, substitutionwiththenaturallyoccurringcytosineanalogue,5-methylcytosine, shifts the preferred rotational position by 3 bp, whereas incorporating 2,6-diaminopurine shifts it 2 bp in the opposite direction. These manipulations potentially would alter the accessibility of a protein recognition sequence on the surface of the histone octamer. We propose that exocyclic groups impose steric constraints on protein-induced DNA wrapping and are also important in determining the orientation of DNA on a protein surface. In addition, we consider the implications of the selection of A-T and G-C base pairs in natural DNA.The local deformation of DNA by DNA-bending proteins can substantially exceed the normal conformational fluctuations of DNA free in solution. Although the interactions that determine the local sequence-dependent conformation of free DNA are relatively well understood (1-7), the nature of the chemical constraints that limit its further deformation have not been explored extensively. The nucleosome core particle in which the central 125 bp of DNA are wrapped in 1.6 superhelical turns about the histone octamer (8, 9) provides a good system to study this problem. In this particle both the major and minor grooves are compressed on the inside of the wrapped DNA and widened on the outside. A major determinant of the rotational placement of the DNA in the nucleosome is the periodic occurrence of short A͞T-rich sequences in helical phase and of short G͞C-rich sequences in the opposite phase. These short sequences favor local conformations with narrow and wide minor grooves, respectively, and thus together facilitate the tight wrapping of DNA (10-16).To investigate the nature of other chemical determinants that constrain the wrapping of DNA around the histone octamer we chose a prokaryotic DNA sequence derived from the Escherichia coli tyrT promoter. Previous studies have shown that the histone octamer occupies a preferred rotational position on this DNA (10). We modified this DNA by replacing the naturally occurring bases with analogues either lacking or containing additional exocyclic groups b...

Genome-wide nucleosome map and cytosine methylation levels of an ancient human genome

“…Intriguingly, the zone depleted for methylation is enriched in CpG dinucleotides (and strong dinucleotides in general, CC, CG, GC, GG; position À30 to +30), suggesting that the presence of nucleosomes protects DNA from methylation in vivo, in agreement with recent experimental results in HeLa cells (Felle et al 2011). Additionally, the drop in methylation levels could ease steric constraints while wrapping DNA around the nucleosome, consistent with the known reduction in CpG deformability following methylation (Perez et al 2012) and the relatively increased difficulty to assemble nucleosomes in vitro on methylated templates compared to methylation-free templates (Buttinelli et al 1998). …”

“…In particular the strandspecific purine versus pyrimidine patterns, which are reversecomplemented at the dyad, would contribute strongly to this. Furthermore, the observed hypomethylation of DNA spanning nucleosome cores might also partake in nucleosome positioning by increasing DNA deformability (Buttinelli et al 1998;Perez et al 2012). All in all, our findings illustrate how aDNA can offer an original source of information to advance our understanding of nucleosome biology.…”

Hepatitis Delta Antigen Requires a Flexible Quasi-Double-Stranded RNA Structure To Bind and Condense Hepatitis Delta Virus RNA in a Ribonucleoprotein Complex

“…The dsDNA binding activities of histones HMG-D and FIS have also been shown to be strengthened by incorporation of inosine, which decreases the stability of base pairs and increases flexibility of dsDNA, and weakened by diaminopurine, which increases the stability of the helix and decreases dsDNA flexibility (41,42,46,47). Therefore, we examined the effects of incorporating inosine or diaminopurine on the binding of HDAg-160 to ); the region mutated is enlarged.…”

Accommodation and Repair of a UV Photoproduct in DNA at Different Rotational Settings on the Nucleosome Surface

“…Indeed, the crystal structure of a CTD site shows that disruption in DNA structure is localized within three bps, two of which are the TT dimer (22). Interestingly, changing DNA flexibility by changing all exo-cyclic groups in the minor or the major grove can increase ⌬⌬G of DNA-histone interactions by as much as 11 kJ/mol, or an average 0.04 kJ/mol/nucleotide (42). In this context, 0.6 -1.1 kJ/mol observed for one CTD lesion is a big change in free energy for one site.…”

“…Precisely, histone-DNA interactions in nucleosomes lead to compression forces in the major and the minor groove that depend on DNA sequence, because A:T, G:C, or G:5-me C base pairs have different groups in each groove (42). The TG motif (A/T) 3 NN(G/C) 3 NN is uniquely suited to accommodate these compression forces (33), which leads to a very specific rotational setting on the nucleosome surface that is stabilized by an additional 2-4 kJ/mol of favorable free energy relative to bulk genomic DNA (36).…”